Project Summary A hallmark of human cancers is their ability to avoid replicative senescence and achieve cellular immortality. Over 90% of cancers achieve this through re-expression of telomerease reverse transcriptase (TERT). TERT encodes the enzymatic component of the ribonucleoprotein complex telomerase, which extends chromosome ends by adding repetitive sequences called telomeres. TERT is transcriptionally silenced in somatic cells, but remains active in stem cells. Recently, non-coding mutations in the promoter of TERT were described in many cancers, including 83% of glioblastoma (GBM), the most common and deadly adult brain tumor, and 80% of oligodendroglioma. TERT promoter mutations are now considered the third most common mutations in human cancer. These mutations serve to reactivate TERT through creation of a canonical E26 transformation specific (ETS) binding motif for the transcription factor GA binding protein (GABP), which has been demonstrated by our lab to be a necessary factor in TERT reactivation and maintaining cellular immortality. It is unclear if genetic alterations in oncogenic signaling pathways cooperate with TERT promoter mutations to fully reactivate telomerase. Depleting the tetrameric subunit of GABP has been demonstrated to reverse cellular immortality in GBM. However, transcription factors are notoriously difficult to target therapeutically and therefore additional means of reducing GABP transcriptional activity at TERT are needed. Transcription of GABP and other ETS factors is regulated by upstream signaling, and these pathways could be targeted as a means of inhibiting TERT. Thus far, no upstream signaling pathways controlling TERT promoter activity have been described in glioma. Preliminary evidence has revealed two candidate pathways altered in glioma?the tumor-promoting AMPK and the tumor-suppressive CIC, that regulate the mutant TERT promoter. This study aims to delineate the role of this pathways in telomere maintenance and cellular immortality. This research will deepen our understanding of the functional role of non-coding mutations and GABP transcriptional regulation in cancer, and will provide insights into different means of targeting GABP to reverse cellular immortality in GBM. Telomerase has long been an attractive therapeutic option, but attempts to target telomerase have been largely unsuccessful, at least in part due to depletion of TERT expressing stem cells. Understanding how tumor suppressive and tumor promoting signaling pathways coordinate to activate the TERT promoter in a GABP-dependent manner would provide novel therapeutic avenues for reversing cellular immortality in adult glioma in a cancer-specific manner.